To develop a self-report Chinese version of the Yale-Brown Obsessive–Compulsive Scale Modified for Body Dysmorphic Disorder (BDD-YBOCS) and determine its validity and reliability in patients seeking a consultation with a plastic surgeon or undergoing plastic surgery in China.t test and bivariate Pearson test. Content validity was established through expert interviews. Construct validity was measured with exploratory factor analysis (EFA) and confirmatory factor analysis (CFA). Convergent validity and discriminant validity were analyzed using Pearson’s correlation to evaluate the association between the self-report Chinese version of the BDD-YBOCS and the Body Dysmorphic Disorder Questionnaire (BDDQ). Internal reliability was assessed using Cronbach’s α.Forward and backward translation and cultural adaptation of the BDD-YBOCS were performed according to recommended guidelines. The self-report Chinese version of the BDD-YBOCS was psychometrically tested using data collected from a cross-sectional validation study, which included 240 patients seeking a consultation with a plastic surgeon or undergoing plastic surgery at the Department of Plastic Surgery, Jiangsu Province Hospital of Chinese Medicine, Nanjing, China, between May and September 2020. Item analysis used the independent sample χ2/df = 1.322, RMSEA = 0.054, GFI = 0.904, NFI = 0.902, CFI = 0.974 and TLI = 0.966). The scale had good convergent and discriminant validity. Cronbach’s α for the scale was 0.871 (P < 0.001) and ranged from 0.852 to 0.873 when individual items were removed.No items were removed from the original English version of the BDD-YBOCS based on expert interviews and factor analysis. A total of 220 patients completed the study survey (91.7%). EFA extracted 3 factors, which accounted for 64.50% of the variance. CFA supported a 3-factor structure (The self-report Chinese version of the BDD-YBOCS shows good validity and reliability for use in patients seeking a consultation with a plastic surgeon or undergoing plastic surgery in China.www.springer.com/00266.This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors  According to the Diagnostic and Statistical Manual of Mental Disorders V (DSM-V) , body dyn = 42/2552) (95% CI 1.2–2.1%), and higher in women (1.9%) than men (1.4%) [The prevalence of BDD in the general population is estimated at 0.7–13% . Specifin (1.4%) . Previoun (1.4%) , 12. A mn (1.4%) . It is eThe Yale-Brown Obsessive–Compulsive Scale Modified for Body Dysmorphic Disorder (BDD-YBOCS) is a 12-item, semi-structured, rater-administered measure that assesses BDD severity during the past week . The scaThe validity and reliability of the BDD-YBOCS in patients undergoing plastic surgery in China have not been assessed, and currently, there is no screening tool for BDD in China. The objective of this study was to translate and culturally adapt the BDD-YBOCS to a self-report Chinese version and determine its validity and reliability in patients seeking a consultation with a plastic surgeon or undergoing plastic surgery in China.Patients seeking a consultation with a plastic surgeon or undergoing plastic surgery at the Department of Plastic Surgery, Jiangsu Province Hospital of Chinese Medicine, Nanjing, China, between May and September 2020, were eligible for this study. Inclusion criteria were: (1) age > 18 years; (2) attending our hospital seeking a consultation with a plastic surgeon or undergoing plastic surgery; (3) voluntarily willing to participate in the study; and (4) able to complete the study survey. Exclusion criteria were (1) inability to understand the study survey; (2) severe physical deformities resulting from tumors or other conditions; or (3) mental disorders such as schizophrenia, depression, or phobia.The study was conducted according to the ethical standards of the institutional and national research committees and the 1964 Declaration of Helsinki and its later amendments or comparable ethical standards. All patients provided written informed consent to participate in the study. Researchers were trained on how to administer the questionnaires and collect data.The BDD-YBOCS is a 12-item semi-structured clinician-rated scale designed to measure BDD symptom severity. The first five items assess excessive obsessional preoccupations with perceived appearance defects, including time preoccupied, interference and distress due to the preoccupations, resistance against the preoccupations, and control over the preoccupations. Items 6 through 10 assess compulsive behaviors, such as excessive grooming and mirror checking. Items 11 and 12 assess insight into appearance beliefs and avoidance due to BDD symptoms, respectively. Each item is scored on a 5-point scale, ranging from 0 (not affected) to 4 (extremely affected). A total score is calculated as the sum of the scores for each individual item, for a maximum of 48. The BDD-YBOCS has demonstrated good validity and reliability among patients undergoing cosmetic or plastic surgery and is the most widely used measure in studies evaluating the efficacy of treatments for BDD .n = 222). It exhibited a sensitivity of 100% and a specificity of 93%, and the correlation coefficient of each item was between 0.808 and 1.000 (P < 0.001). This suggests the Chinese translation of the BDDQ has high validity and reliability [The BDDQ is a brief 5-item self-administered questionnaire based on the DSM-IV diagnostic criteria for BDD . The firiability .Forward and backward translation and cultural adaptation of the BDD-YBOCS were performed according to recommended guidelines to develop a self-report Chinese version of the BDD-YBOCS .Two independent bilingual translators with Master’s degrees and experience in psychology or plastic surgery created Chinese translations of the BDD-YBOCS and consolidated them into a single version after discussion. Two independent bilingual native Chinese translators, one with a doctorate in statistics from the USA and the other a professional English translator, back-translated the consolidated version from Chinese to English. The translated and original versions were reviewed to ensure translated items retained the meaning of the original items, without confusion.“Is it possible that your defect might be less noticeable or less unattractive than you think it is?” scored as 0=excellent insight, 1=good insight, 2=fair insight, 3=poor insight, and 4=absent insight); therefore, the anchors were replaced with: 0=yes, very much possible, 1=possibly yes, 2=not sure, 3=possibly not, 4=no, definitely not possible.The Chinese translation of the BDD-YBOCS was revised into a self-report scale by two professors from the Psychology Department, who modified wording and provided notes. The revised scale retained the 12 items included in the original scale, but items were reordered. General information on the respondents was collected, including gender, age, marital status, highest education, occupation, and history of plastic surgery. An expert committee comprised of 10 native Chinese-speaking experts in the fields of plastic surgery and psychology evaluated the cultural aspects and language of the self-report Chinese version of the BDD-YBOCS and proposed modifications. Items marked as “unclear” by over 20% of the committee participants were revised. The clarity of the items in a pre-final version was pilot tested in 20 patients. Of these, 8 patients misunderstood item 11 and its anchors  and confirmatory factor analysis (CFA). Convergent validity and discriminant validity were analyzed using Pearson’s correlation to evaluate the association between the total score on the self-report Chinese version of the BDD-YBOCS and the BDDQ. Internal reliability was assessed using Cronbach’s α.Item analysis used the independent sample A total of 240 patients seeking a consultation with a plastic surgeon or undergoing plastic surgery were invited to participate in this study. Of these, 12 patients declined to join the study, and 8 patients did not complete the study survey. Finally, data from 220 (91.7%) patients were included in the analyses.The sociodemographic and clinical characteristics of the included patients are summarized in Table t test, the differences between the scores for each item in the upper and lower groups were significantly different (P < 0.01), indicating all items provide good discrimination. The bivariate Pearson test showed a significant positive correlation between the score for each item and the total score for the self-report Chinese version of the BDD-YBOCS , indicating all items are closely associated with the scale.Item analysis computed the critical ratio and the item-total correlation. Respondents with the highest and lowest scores on the self-report Chinese version of the BDD-YBOCS were separated into upper and lower groups comprised of the top 27% and bottom 27% of the scores. Mean total score of the upper group was 18.63 (SD = 1.67), and mean total score of the lower group was 4.1 (SD = 0.35). On the independent sample r = 0.557, P < 0.01), indicating good criterion validity.Ten experts were invited to independently assess the content validity of the self-report Chinese version of the BDD-YBOCS. The item content validity index (ICVI) was 0.90 (criteria: > 0.78), indicating that the scale accurately represents all facets of the construct BDD. The total score of the self-report Chinese version of the BDD-YBOCS was significantly correlated with the total score of the BDDQ (P < 0.05). Results indicated that the sample size (n = 110) was adequate .Before performing factor analysis, the Kaiser–Meyer–Olkin test (KMO) and Bartlett’s sphericity test were conducted to measure sampling adequacy . Factor 1, “obsessions,” was strongly associated with items 1, 2, 3, 4, and 5 (loadings ranged from 0.642 to 0.776) and accounted for 28.698% of the variance. Factor 2, “compulsions,” was strongly associated with items 6, 7, 8, 9, and 10 (loadings ranged from 0.551 to 0.793), and accounted for 21.463% of the variance. Factor 3, “additional,” was strongly associated with items 11 and 12 (loadings 0.809 and 0.596), and accounted for 14.335% of the variance. All factor loadings were > 0.50, suggesting that the three factors are related and are elements of the construct BDD (Table n = 110) supported a 3-factor structure for the self-report Chinese version of the BDD-YBOCS, with factor loadings ranging from 0.54 to 0.83. Model fit indices were χ2/df = 1.322 (criteria: < 3), CFI = 0.974, GFI = 0.904, NFI = 0.902, TLI = 0.966  and RMSEA=0.054 (criteria: < 0.08) (Table CFA in an independent sample (Composite reliability (CR) considering factor loadings was used to assess internal consistency in scale items. A higher CR value represents a higher internal consistency of the factor, with 0.7 as the acceptable threshold. The CR of each factor was greater than 0.776 Table .Table 4FConvergent validity was assessed using the average variance extracted (AVE) (criterion > 0.5). AVE values for the factors demonstrated acceptable convergent validity Table .Discriminant validity was assessed by comparing the square root of AVE of each factor with correlations between pairs of factors (Fornell and Larcker 1981). The square root of AVE was larger than the correlation between any pair of factors representing satisfactory discriminant validity Table .Table 5DCronbach’s α for the self-report Chinese version of the BDD-YBOCS and the obsessions, compulsions, and additional factors were 0.871, 0.824, 0.797, and 0.637 (criteria: Cronbach’s α > 0.70), respectively, with Cronbach’s α ranging from 0.852 to 0.873 when individual items were deleted. The deletion of any item did not increase the Cronbach’s α sufficiently to imply the self-report Chinese version of the BDD-YBOCS is better without that item. Corrected item-total correlations (ITC) ranged from 0.363 to 0.707  and the item-total correlation . Results confirmed that no items are redundant and should be removed from the tool. In pilot testing, most patients understood the items and answered all items within 5–10 minutes. The anchors for item 11 “Is it possible that your defect might be less noticeable or less unattractive than you think it is?”, which assesses insight and distress, were amended to measure “self-belief” and be applicable to a self-report format.Item analysis of the self-report Chinese version of the BDD-YBOCS involved calculating the critical ratio  observed between the self-report Chinese version of the BDD-YBOCS and the BDDQ indicates that both instruments can identify the obsessive and compulsive behaviors that characterize BDD.The ICVI of the self-report Chinese version of the BDD-YBOCS was 0.90 (criteria: > 0.78), indicating that the scale has good content validity. To date, the BDDQ is the only instrument that has been translated and validated to measure dissatisfaction with physical features and facilitate screening and diagnosis of BDD in China . The strFactor analysis is a standard method to evaluate construct validity and test whether a scale effectively measures a construct of interest. EFA and CFA are two frequently used forms of factor analysis. Both are based on general factor models, but have differences in methodology and scale evaluation. Psychometric testing of the original version of the BDD-YBOCS included EFA, which showed three factors  accounted for 59.6% of the variance . Furtherα = 0.871), and the factors obsessions, compulsions, and additional yielded similar scores .Internal consistency reliability verified that the self-report Chinese version of the BDD-YBOCS addresses the construct of BDD (Cronbach’s The present study had some limitations. First, psychometric testing of the self-report Chinese version of the BDD-YBOCS was performed using data collected from patients seeking a consultation with a plastic surgeon or undergoing plastic surgery at one institution, leading to the potential for selection bias. Future studies require larger sample sizes across multiple institutions to ensure data are generalizable to the population in China. Second, we developed a self-report version of the BDD-YBOCS. Self-report measures are limited by response biases and constraints on self-knowledge. We recommend that our version of the BDD-YBOCS is used for preliminary screening of possible cases of BDD. Third, our evaluation of the criterion validity of the self-report Chinese version of the BDD-YBOCS was restricted to a comparison with one tool, the BDDQ. Fourth, plastic surgeons used subjective judgment to determine whether patients met the exclusion criterion for a pre-existing mental disorder. Patients that were considered “not quite right” were referred to the psychiatric department for further evaluation. This method of identifying pre-existing mental disorders may have caused some potentially eligible patients to be excluded from the study. Last, we did not determine the sensitivity and specificity of the self-report Chinese version of the BDD-YBOCS or identify cutoff points for symptom severity.The current study suggests that the self-report Chinese version of the BDD-YBOCS is a valid and reliable instrument to assess BDD in patients seeking a consultation with a plastic surgeon or undergoing plastic surgery in China. It may help researchers screen and evaluate interventions aimed at reducing the symptoms of BDD. Future research is needed to further evaluate the instrument and support its application in China.


Chronic liver diseases including hepatocellular carcinoma (HCC) create a state of chronic inflammation that affects the brain via the liver–brain axis leading to an alteration of neurotransmission and cognition. However, little is known about the effects of HCC on the hippocampus, the key brain region for learning and memory. Moreover, radiotherapy used to treat HCC has severe side effects that impair patients’ life quality. Thus, designing optimal strategies, such as chronotherapy, to enhance the efficacy and reduce the side effects of HCC treatment is critically important. We addressed the effects of HCC and the timed administration of radiotherapy in mice on the expression of pro-inflammatory cytokines, clock genes, markers for glial activation, oxidative stress, neuronal activity and proliferation in the hippocampal neurogenic niche. Our data showed that HCC induced the upregulation of genes encoding for pro-inflammatory cytokines, altered clock gene expressions and reduced proliferation in the hippocampus. Radiotherapy, in particular when applied during the light/inactive phase enhanced all these effects in addition to glial activation, increased oxidative stress, decreased neuronal activity and increased levels of phospho(p)-ERK. Our results suggested an interaction of the circadian molecular clockwork and the brain’s innate immune system as key players in liver–brain crosstalk in HCC and that radiotherapy when applied during the light/inactive phase induced the most profound alterations in the hippocampus. Hepatocellular carcinoma (HCC) represents the most prevalent primary hepatic malignancy and the third leading cause of cancer deaths with an 8.3% mortality rate . ImportaHCC pathogenesis includes inflammatory damage of hepatocytes and regeneration leading to fibrotic deposition, distorted hepatic architecture and finally tumor development associated with liver function impairment . RecentlThe circadian system in mammals is a hierarchically organized internal time-keeping system which coordinates rhythms of physiology and behavior with the 24h solar day . The hypIn this study with a mouse model, we analyzed the effect of HCC and the timed application of radiotherapy on the expression of pro-inflammatory cytokines, clock genes, markers for glial activation, oxidative stress, neuronal activity and neural progenitor cell proliferation in the hippocampal neurogenic niche, a key structure for neural plasticity and cognition. Additionally, we investigated the protein levels of phosphorylated extracellular signal-regulated kinase 1/2 (ERK1/2), an essential modulator of hippocampal functions . Chronic treatment with phenobarbital (PB) that was used here to promote HCC development affected the expression of pro-inflammatory cytokines and clock genes as well as microglia activation in the hippocampus. HCC resulted in an upregulation of the expression of pro-inflammatory cytokines, a downregulation of neural progenitor cell proliferation and p-ERK protein levels as well as changes in clock gene expression in the hippocampus. Radiotherapy, in particular when applied during the light/inactive phase of mice, enhanced all of these effects and resulted in glia activation, increased oxidative stress and changes in neuronal activity in the hippocampus. ad libitum) in temperature-controlled rooms under normal light–dark cycles 00, light off 06:00 pm, ZT12). Experiments during the dark phase were performed under dim red light.Male mice (PER2::LUC with C57BL/6J background) were used ,17. MiceDetails on the treatment of the mice are described in . Brieflyn = 3 for each time point). Prior to irradiation, the mice were anesthetized by i.p. injection of a mixture of ketamine/xylazine (100/10 mg/kg) and then fixed with an adhesive tape so that their abdomen was facing the irradiation source. Mice were exposed to a dose of 10 Gray (Gy) for 10 min at (175 kV and 15 mA) using a Gulmay RS225 X-ray system . A total of 10 Gy was used according to the dose applied in human palliative radiotherapy [n = 3 mice for each time point, to keep the scarification of all mice close to each other in every time point. Only animals with a similar tumor size were included for further analyses; animals without tumors or with smaller or larger tumors were excluded.Details on the irradiation of the mice are described in . BrieflyIL-1a, IL-1b, IL-1r1, IL-6 and TNFα, as well as the expression of clock genes, Clock, Bmal1, Per1, Per2, Cry1, Cry2 and Rev-Erba were analyzed. The sequences of the used primers are listed in Mice from each group were killed at four different time points: ZT02, ZT08, ZT14 and ZT20, 48 h after irradiation/handling. The entire hippocampus was dissected, snap frozen and then stored at −80 °C. RNA was isolated using RNeasy Plus Universal Mini Kit  according to the manufacturer’s instructions. The concentration of RNA in the samples was analyzed using a NanoDrop 2000 spectrophotometer  and its purity was assessed by the ratio of absorbance at 260 nm and 280 nm. A Revert Aid First Strand cDNA Synthesis Kit  was utilized to prepare cDNA using 1 µg RNA. Negative controls were used to exclude the DNA contamination. The expression levels of several genes encoding for pro-inflammatory cytokines, including IL-1a, IL-1b, IL-1r1, IL-6 and clock genes Clock, Bmal1, Per1, Per2, Cry1, Cry2 and Rev-Erba, were normalized to housekeeping genes using the Pfaffl method [Gapdh and Rn18s or ß-Actin, Gapdh and Rn18s were used as housekeeping genes for the relative expression levels of pro-inflammatory cytokines and clock genes, respectively. The quality of the PCR amplification product was confirmed by melting curves and agarose gel electrophoreses. The gene accession number was obtained from the NCBI database (Gene database/GenBanke) (https://www.ncbi.nlm.nih.gov/gene (accessed on 16 December 2022)). Amplicon size for the target genes was detected using Primer-BLAST (https://www.ncbi.nlm.nih.gov/tools/primer-blast/index.cg (accessed on 16 December 2022)). Real-time qPCR was performed using 1 ng cDNA and SYBR GREEN , in an ABI StepOne Plus Real-Time PCR System  with the following program: activation at 95 °C for 5 min, then denaturation with 40 cycles at 95 °C for 3 s and amplification and annealing at 60 °C for 20 s. A standard curve for each primer was used to calculate primer efficiency. mRNA expression levels of pro-inflammatory cytokines l method  to obtaiMice from each group were sacrificed 48 h after irradiation/handling at four different time points: ZT02, ZT08, ZT14 and ZT20 via i.p. injection of ketamine/xylazine (100/10 mg/kg) then transcardially perfused with 0.9% NaCl followed by 4% formalin.The brains were removed from the skull and post-fixed in 4% formalin for 24 h. Brains were cryoprotected in 20% sucrose, followed by 30% sucrose. Brains were cut on a Leica CM3050 cryostat  into 30 µm thick coronal sections along the rostro-caudal extent of the hippocampus. Primary and secondary antibodies used for immunohistochemistry are listed in 2O2 for 30 min at room temperature (RT). To prevent non-specific staining, sections were pre-incubated in a blocking buffer of 5% normal goat serum in PBS-Triton 0.2% (PBST) for one hour at RT. Then, sections were incubated overnight at 4 °C with antibodies against markers for neuronal activity (c-FOS) or proliferation (Ki67). The sections were then washed in PBST and incubated with a biotin-conjugated secondary antibody for one hour at RT. The sections were then rinsed with PBST and then treated with the Vectastain Elite ABC kit  for one hour at RT. This was followed by rinsing with PBST and then by incubation in 0.05% 3,3′-diaminobenzidine  for 5 min at RT. Sections were finally washed with PBS and coverslipped with DePeX .For chromogenic immunohistochemistry, sections were washed in phosphate-buffered saline (PBS) and incubated with 0.6% HFor immunofluorescence, sections were washed and incubated with a blocking buffer for one hour. The sections were incubated overnight at 4 °C with antibodies against markers for microglial cells IBA1, astrocytes GFAP or the oxidative stress 8-hydroxy-2′-deoxyguanosine (8-OHdG) , and the3. Immunofluorescent signals (IR) for IBA1, GFAP and 8-OHdG were recorded using 20X objectives and respective filters of KEYENCE BZ-900E fluorescent microscope. The mean intensity in the hippocampus was quantitatively analyzed above the background intensity and expressed as arbitrary units (A.U.) using Image J software ) as previously described [Prior to image acquisition, the experimental condition was obscured to the investigator. Images were acquired using a BZ-900E microscope . The microscope settings were kept constant for all images of one staining. c-FOS immunoreactive (+) and Ki67+ cells were recorded in the bright field mode with 40X objective. Immunoreactive cells were counted in a delineated area of the hippocampal dentate gyrus (DG) using BZ-II analyzer software ; the mean cell density was calculated and displayed as number of cells/mmescribed .® Evolution tissue homogenizer . A BCA kit  was used to determine the protein concentration. Gel electrophoresis of independent samples and immunoblotting were performed using Novex XCell Sure Lock system and PVDF membranes as described [The mice were sacrificed 48 h after irradiation/handling at four different time points: ZT02, ZT08, ZT14 and ZT20 and the hippocampus was dissected and homogenized in RIPA buffer containing HALT proteinase and phosphatase inhibitor  using a PRECELLYSescribed . Membrant-tests were used to analyze significant differences between two groups at a specific time point, while the comparison between different time points within one group was performed using ordinary one-way analysis of variance (ANOVA) followed by Tukey’s post-hoc tests. When the data violated the normal distribution, the non-parametric Mann–Whitney-U test was used to compare two groups at a specific time point and Kruskal–Wallis followed by Dunn’s multiple comparisons test were applied to detect differences among multiple time points within one group. Data are expressed as mean ± standard error of the mean (SEM). The results were regarded as statistically significant if p < 0.05.GraphPad Prism 8 software  was used for statistical analysis. Anderson–Darling, Kolmogorov–Smirnov and D’Agostino and Pearson tests were used to test the normality of the data. When the data were normally distributed, multiple IL-1a, IL-1b and TNF-α during the light/inactive phase, which were blunted in control-PB mice (IL-1r1 at ZT02 (p = 0.04) and ZT14 (p = 0.002) and of TNF-α at all-time points  than control-PB mice  as compared to non-irradiated HCC-bearing mice , while irradiation at any other time point had no significant effect (In the granule cell layer (GCL) of the hippocampal dentate gyrus (DG), the number of c-FOS+ cells, a marker of neuronal activity, was not significantly different between untreated and PB-treated mice . HCC hadt effect A,C. The number of the proliferating Ki67+ cells in the neurogenic niche within the DG was not different between untreated and PB-treated mice . HCC resClock, Bmal1, Per1, Cry1 and Cry2 mRNA expression levels showed a time-of-day-dependent variation with a peak at ZT02 while Per2 and Rev-Erba did not show a time-of-day-dependent variation  (The relative protein levels of p-ERK in the hippocampus were not significantly different between PB-control mice and HCC-mice. Irradiation at the late light/inactive phase (ZT08) led to a significant increase in the protein levels of p-ERK compared to the HCC-mice ( = 0.02) . IrradiaHere, we have shown that HCC resulted in an increased expression of pro-inflammatory cytokines and decreased proliferation and changes in clock gene expression in the hippocampus. Irradiation, in particular when applied in light/rest phase, enhanced all of these effects and also caused the activation of glia, oxidative stress and increased p-ERK protein levels while cFos-IR was decreased.Per2 and Rev-Erba showed a peak during the early light/inactive phase in agreement with previously reported findings [IL-1a, IL-1b and TNF-α showed the same peak, suggesting an interaction of the molecular clockwork and the innate immune system in the hippocampus. There is increasing evidence of an interaction between the molecular clockwork and neuroinflammation [Bmal1 [Rev-Erba [The hippocampal-dependent cognitive functions such as memory formation and extinction are largely time-of-day-dependent and are influenced by the circadian system . In the findings ,29. Inteammation ,31. In an [Bmal1  and Rev-Rev-Erba  with theRev-Erba . SurprisRev-Erba ,35,36,37In mice with HCC, the expression of pro-inflammatory cytokines and clock gene expression was increased in the hippocampus, indicative of a liver–brain crosstalk. Similarly, a recent study showed that circadian disruption enhanced breast cancer-mediated inflammation in the brain . MoreoveClock, Cry1, Cry2, Rev-Erba and the pro-inflammatory cytokines IL-1b, TNF-α, oxidative stress and the activation of microglia and astrocytes. Clock genes are important players in microglial activation and pro-inflammatory responses [Radiotherapy is known to induce pro-inflammatory responses and microglial activation in the hippocampus , astroglesponses ,57. Inteesponses . In contesponses  and are esponses . The extesponses . It has esponses . MAPK/EResponses . Additioesponses . The MAPesponses . IrradiaIn conclusion, our study supports the hypothesis of an interaction of the molecular clockwork and the innate immune system and that pro-inflammatory cytokines are key players in liver–brain crosstalk in the context of liver cancer and thus, might help to refine therapeutic targets for the amelioration of central symptoms. Most importantly, we showed that radiotherapy induced the most profound effects on the expression of pro-inflammatory cytokines and clock genes, glial activation, oxidative stress and neuronal activity in the hippocampus when applied during the light/inactive phase, presumably via an ERK pathway-related mechanism. Thus, radiotherapy application during the (late) active phase not only has the highest efficacy on tumor tissue and the least side effects on liver tissue as we showed earlier but also the least side effects on the hippocampus. However, translational studies are needed to prove this hypothesis in human patients.


The full-field ERG is useful for index rod- or cone-mediated retinal function in rodent models of retinal degeneration. However, the relationship between the ERG response amplitudes and visually guided behavior, such as flicker detection, is not well understood. A comparison of ERG to behavioral responses in a light-damage model of retinal degeneration allows us to better understand the functional implications of electrophysiological changes. Flicker-ERG and behavioral responses to flicker were used to determine critical flicker frequency (CFF) under scotopic and photopic conditions before and up to 90 d after a 10-day period of low-intensity light damage. Dark- and light-adapted ERG flash responses were significantly reduced after light damage. The a-wave was permanently reduced, while the b-wave amplitude recovered over three weeks after light damage. There was a small, but significant dip in scotopic ERG CFF. Photopic behavioral CFF was slightly lower following light damage. The recovery of the b-wave amplitude and flicker sensitivity demonstrates the plasticity of retinal circuits following photopic injury. The full-field electroretinogram (ERG) is commonly used to quantify retinal function in humans  as well Continuous exposure to low-intensity light causes photoreceptor apoptosis in albino animals ,10,11. UHowever, the characterization of rods as more vulnerable to light damage is not consistently reported. Sugawara, Sieving and Bush showed reductions in the dark- and light-adapted ERG in rats exposed to 1000–3000 lux for up to 48 h . SensitiIn the present study, rod and cone function was monitored for 90 days after 10 days of exposure to 280-lux fluorescent light. ERG responses were measured by dark- and light-adapted flashes and sinusoidal flicker. Psychophysical measures of CFF were conducted in a water maze. The differences between the effect of light-induced retinal degeneration on threshold measures versus maximum responses were observed. Strong parallels were noted between behavior and ERG measures of flicker threshold. This light-damage model demonstrates significant recovery of the flash ERG, however, CFF thresholds were minimally disturbed by a significant loss of photoreceptors.max) of the a- and b-wave of the flash response were lower across all experimental days following light damage. max values measured before light damage and on recovery day (R) 6, 20, 48, and 90 in the groups of control (filled circles) and test animals (open squares). In max responses to light-damaged animals (n = 9) were a third lower than the pre-light damage responses (p < 0.05). By R20, the b-wave recovered over 80% of its original amplitude, whereas the loss of the a-wave was permanent (p < 0.05). p < 0.001). However, there was no significant difference between the a-wave amplitudes of the light-damage and control groups at R90. Interestingly, b-wave amplitudes were affected very little by the loss of a-waves, there was no decrement in control animals and in addition, the 33% loss of a-waves in the LD group results in only 19% decrement of the b-waves in these animals at R90. Light damage (LD) treatment led to ERG amplitude reduction. Dark-adapted maximum responses  and steadily declined thereafter. By R90, the a-wave had declined by 36% compared to the Pre-LD values. In contrast, the b-wave showed an initial decline of 30% but thereafter recovered, resulting in only a 10–15% loss of amplitude.Cone ERG responses to bright flashes were measured on a 505 nm rod-saturating background c,d. Rmax1/2, photons/µm2 at cornea) of the a- and b-waves. 1/2 of a- and b-waves at Pre-LD, R6, R20, and R34. At R6, a half-maximal a-wave could be elicited with a 1.7-fold less light than pre-light damage (p < 0.05). However, the a-wave sensitivity was not significantly different at R20 and thereafter. Thus, after following the first cohort of animals for 90 days post light damage, the second litter of animals was only followed for 34 days after the end of the light-damage regimen  than behavioral CFF under scotopic conditions. Light-damaged animals did not exhibit significant behavioral differences under scotopic conditions and for ERG measures, only results from day R6 showed significant scotopic CFF loss (p < 0.01).p < 0.001) over the entire recovery period between R6 and R48 (p < 0.05) and photopic (p < 0.01) conditions. Thus, the signals we measure with flicker-ERG do not match the behavioral measures of flicker sensitivity in our forced-choice swimming test. Under photopic conditions, the initial ERG CFF was higher (47.8 Hz) than behavioral CFF (40.8 Hz). Behavioral CFF for photopic lighting was significantly depressed  and light-damaged retinas . The calculated area of the ONL for a section of the retina from light-damaged animals was 324 µm2, significantly less (p < 0.01) than the matched area of ONL in the control animals (417 µM2). INL areas were also compared in light-damaged animals to controls, however, the difference was not statistically significant. The measures of ONL thickness matched the same comparison made by ONL nuclei counts (Photoreceptor loss was quantified by comparing the outer (ONL) and inner (INL) nuclear layer thickness of light-damaged versus control animals . The outi counts .Albino rats exposed to low-intensity light for 10 days showed transient changes in b-wave amplitudes, a-wave sensitivity, and CFF values. These changes were markedly different from the results obtained previously, detailing the permanent loss of the rod- and cone-driven ERG flicker signal in aging RCS rats  and moreIn the present study, we measured a 33% loss of dark-adapted b-wave amplitude six days after light damage. At the end of our study, ONL thickness showed a persistent 22% loss, but the b-wave had recovered. Other investigators have noted b-wave recovery after photoreceptor loss ,23,24 anCngb1-x1) [We previously tested the flicker function in a ß subunit knockout (KO) mouse model (ngb1-x1) . In thisngb1-x1) . FurtherA 3 µV criterion of response was used for determining ERG CFF since it is near the root mean squared noise in our averaged recordings. Other studies have also used a 3 µV criterion amplitude for flicker ERG analysis ,35. InitA single study by Coile et al., reported behavioral CFF exceeding ERG CFF measures in dogs . HoweverThere was a significant dip in scotopic ERG CFF after light damage at R6. This temporary deficit may be the result of photostasis , in whicAfter light damage, there was a small decrease in the light-adapted a-wave at R6, suggesting an impairment of cone function. Over the time course of these trials, the impairment progressively worsened, such that at R90, a statistically significant difference in the a-wave amplitude was noted. This raises the interesting possibility that cone receptors continue to degenerate after light damage, which may also help explain the apparent greater disparity between the ERG and behavioral criterion. Several studies have documented long-term apoptosis in ONL after exposure to high-intensity light ,43,44,45Our study showed that CFF was less affected by light damage than the ERG b-wave amplitude. The minor changes to CFF demonstrate the preservation of threshold responses that mediate behavior despite significant photoreceptor degeneration. The CFF was not significantly different from control animals at the end of our study. These findings suggest that a level of retinal resilience preserves functional circuits or computations despite the loss of photoreceptors. Because the damage induced by low-level constant light damage is easily controlled, it affords the experimenter the opportunity to test the protective measures given in advance of damage, concurrent with the ongoing physiologic insult or during the recovery period. It is known that retinal damage can induce upregulation of neuroprotective mechanisms and compounds and, that enhancing those reactions, for example, by photobiomodulation, can help to preserve retinal structure and function ,47. In oSprague Dawley albino rats were housed under 12-h light/dark conditions (57–140 lux). Animals were trained on a behavioral task for 65 days prior to light damage. All animals were handled according to the principles of the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research.Details of the ERG recording follow those of Rubin and Kraft . Briefly4 photons µm−2 s−1) incident upon the cornea. The wavelength of the stimuli was chosen to match or be near the optimum wavelength for rat rhodopsin and cone opsin [Stimulus intensity was controlled by calibrated neutral-density filters, and the wavelength was 505 nm . ERG responses were obtained using 3 to 20 repeats of a 10 ms stimulus . The int10 of the response amplitudes for each intensity was plotted and fitted with a line to determine electrophysiological CFF using a 3µV criterion voltage [Sinusoidal flicker was produced by a ferro-electric liquid crystal shutter  driven by a pulse-width modulation paradigm. The average Michelson contrast of the flicker stimulus was 0.86. The stimulus frequencies were: 1, 2, 4, 5, 10, 16, 20, 25, 32, and 40 Hz. The on-transient of the 5 s response was ignored and the final 4.5 s of data were averaged into a one- or two-cycle wave to measure the response amplitude. The log voltage .Behavioral CFF was determined by two alternative forced-choice tests in a water maze ,52. A trThe two displays were constructed by placing a translucent sheet (ca 17 cm square) in front of a bank of green LEDs enclosed in a mylar coated funnel. The opening at the end of the funnel (1 ¾ cm) sat flush against the acrylic sheet. Inserting neutral-density filters between the funnel and acrylic sheet attenuated the light intensity. A specially fabricated circuit drove one display at an adjustable flickering frequency (5–80 Hz) and the other at a steady fixed high frequency (180 Hz). The position (left/right) of the steady vs. flickering displays could be switched, and the two displays were matched in time-averaged luminance.Behavioral testing was accomplished in 3 phases . InitialNext, the rat learned to discriminate between the steady and flickering (~6 Hz) stimuli. The rat was sequestered behind a clear starting gate and then required to locate the platform in front of the correct stimulus before being removed from the maze. If the rat passed the divider towards the flickering stimulus, the trial was recorded as an error. This training phase continued until the rat consistently  selected the steady stimulus.CFF was determined by increasing the temporal frequency following a method of limits staircase procedure . In the 3 µm−2 s−1 photons; photopic 6.38 × 106 µm−2 s−1 photons).The position of the correct stimulus during CFF testing followed a Gellerman series . If an aTransparent cages were split along the long axis with one rat per compartment. Minimum bedding was used in the cages. Water and food were provided ad libitum. A fluorescent light bulb  48 inches, 32 W, 2950 lumens, color rendering 85, correlated color temperature 3000 K) was placed above the animal cages for an average illuminance of 280 + 20 lux . Following the light damage, rats were returned to cyclic (12/12) lighting.2 asphyxiation, and whole eyes were harvested and fixed overnight . The tissue was dehydrated and then embedded in plastic . Radial sections, 5 μM thick, were cut  and collected at 500 µM intervals. Slides were stained with Hematoxylin and Eosin and retinal sections were imaged for measures of outer nuclear layer (ONL) and inner nuclear layer (INL) thickness. Measurements were taken at 450 µM increments, starting from the optic nerve head (ONH) in both inferior and superior hemispheres. Only animals surviving 48 days past light, or sham, damage were considered in the statistical analysis of histological data.Rats were euthanized by COt-test with equal variance assumed. Within-subjects comparisons were made by a paired t-test. Flicker analysis and comparisons between the ERG and behavioral CFF were made by repeated-measures ANOVA. Histological analysis was made by a 2-tailed 2-sample t-test. Equal variance and normal distributions were assumed.The between-group significance for ERG data collected at specific experimental time-points  was calculated by a 2-tailed 2-sample


During the last years several data have been reported in literature investigating its efficacy and safety, as well as defining the dosing and application regiments of BoNT-B in the treatment of hyperhidrosis. Moreover, recent studies have been examining its safety profile, which may be different from those known about BoNT-A. The aim of this review is to provide information about what is currently known about BoNT-B in regards to the treatment of focal hyperhidrosis.Botulinum toxin type B (BoNT-B), known as Myobloc The burden of focal hyperhidrosis is directly related to its social, psychological and occupational repercussions, which make it a distressing and disabling condition for affected patients. Although the use of botulinum toxin type A (BoNT-A) has radically changed the management of patients suffering from axillary and palmar hyperhidrosis, and it has gained a positioning in the guidelines of the International Hyperhidrosis Society, several limiting factors related to duration of benefit, and hand muscle weakening related to motor nervous system involvement mainly following BoNT-A diffusion through thenar and hypothenar hand eminence have driven several authors to investigate the efficacy and safety profile of botulinum toxin type B (BoNT-B) for the treatment of focal hyperhidrosis .BoNT-B is produced as relatively inactive, single polypeptide chains with a molecular mass of about 150 kDa consisting of a heavy (H) chain and a light (L) chain of roughly 100 and 50 kDa, respectively, linked by a disulfide bond . The botThe efficacy of BoNT-B in focal hyperhidrosis is related to its interfering action on neural transmission by blocking the release of acetylcholine. BoNT-B inhibits acetylcholine release at the neuromuscular junction via a three-stage process: (1) H-chain-mediated neurospecific binding of the toxin, (2) internalization of the toxin by receptor-mediated endocytosis, and (3) ATP and pH dependent translocation of the L chain to the neuronal cytosol where it acts as a zinc-dependent endoprotease cleaving polypeptides essential for neurotransmitter release.BoNT-B binds to and cleaves the synaptic Vesicle Associated Membrane Protein  which is a component of the protein complex responsible for docking and fusion of the synaptic vesicle to the presynaptic membrane, a necessary step for neurotransmitter release .BoNT-B is available in a ready-to-use formula that does not require reconstitution. However, many physicians choose to dilute it to reduce the pain on injection. When diluting a vial of BTX-B, it is important to consider the bottles are overfilled and actually contain slightly more toxin than the label states, to account for excess lost in the needle tip. BoNT-B is available in three vial configurations of 2500 U, 5000 U, and 10,000 U, with a composition of 5000 U BoNT-B/mL. BoNT-B vials are stable for 30 months when refrigerated and for 9 months at room temperature .The aim of this narrative review is to provide an update on the efficacy and safety profile of BTX-B in the treatment of focal hyperhidrosis.2) after 3 weeks. The duration of anhidrosis was prolonged for 12 weeks when 15 MU BoNT-B was injected, and for 24 weeks when 125 MU BoNT-B was injected. The size of the anhidrotic area decreased with time, indicating partial recovery at the edges. After 3 weeks, the QSART score had significantly decreased to 18% of the baseline and had decreased to zero in most subjects with doses of 62.5 MU or more. After 12 weeks, the QSART had returned to 91% of the baseline in all but one subject and, after 24 weeks, recovery of sudomotor function was complete. Analysis by iodine-starch staining and QSART indicated that BoNT-B suppresses sudomotor function effectively in a concentration-dependent manner.A proof of efficacy of BoNT-B on sweat production was described in 2003 by Birklein F et al. . The aut®/MyoBloc®) with that of BoNT-A (Botox®). Nine patients (High Dose group-HD) received BoNT-A 100MU unilaterally and BoNT-B 4000 MU contralaterally. Ten patients (Low Dose group-LD) received BoNT-A 100 MU and BoNT-B 2000 MU. All patients were blinded as to which preparation was used in which side. All patients except one reported excellent hyperhidrosis improvement in both axillae. None of the patients had residual symptoms on clinical examination. The duration of hyperhidrosis improvement until first recurrence in the HD group was similar between the BoNT-B and BoNT-A sides. In the LD group, duration was prolonged in the BoNT-A side. There was no difference in the duration of hyperhidrosis improvement between the axillae treated with BoNT-B 4000 MU and BoNT-B 2000 MU. Five out of nine patients in the HD group and seven out of ten patients in the LD group reported more application discomfort in the BoNT-B treated axillae. In six out of nine patients in the HD group, and in six out of ten patients in the LD group, the onset of hyperhidrosis improvement appeared earlier in the BoNT-B treated axillae. One patient in the HD group reported dryness of the mouth and eyes and accommodation difficulties. These authors viewed BoNT-B as a safe and efficient treatment for axillary hyperhidrosis. Doses of BoNT-B 2000 MU per axilla seemed sufficient, indicating a conversion factor between BoNT-A and BoNT-B in the order of 1:20, as the autonomic nervous system seems to be relatively more sensitive to BoNT-B than to BoNT-A compared with the motor system.Results reported in In 2004, Hecht MJ et al.  reported®) for axillary hyperhidrosis. Thirteen patients (22 axillae) were recruited to the study. The hyperhidrotic area was defined using the iodine-starch test then measured and photographed. A total of 5000 MU of BoNT-B was administered subdermally. Patients were reviewed at 4, 8, and 12 weeks to assess the outcome objectively (using hyperhidrotic area measurements and photographs) and subjectively (by assessing sweat production and patient satisfaction). A significant reduction in the hyperhidrotic area at follow-up compared to the baseline was demonstrated. The mean percentage reduction in the hyperhidrotic area was 84, 87, and 81% at 4, 8, and 12 weeks, respectively . Patient satisfaction was 100% throughout. Subjective mean percentage reduction in sweat production was 98, 96, and 90% at 4, 8, and 12 weeks, respectively. Side effects were negligible.In the same year, Nelson L et al.  investig®)  or 0.5 mL vehicle  into bilateral axillae. Participants who received a placebo were rolled over and received BoNT-B at subsequent visits. All participants were followed until sweating returned to baseline levels. According to the participant assessment of axillary hyperhidrosis improvement and quality of life scores and the physician assessment scores, a significant difference was observed in treatment response at day 30 in the participants receiving Myobloc (BoNT-B (Myobloc®)) injections. The duration of action ranged from 2.2 to 8.1 months (mean 5.0 months). The adverse event profile included bruising, flu-like symptoms, and dry eyes. In 2005, Baumann L et al.  reported®) with BoNT-A (Onabotulinum toxin-Botox®) to address the question concerning the effective dose. They performed a bilateral paired, single-blinded, randomized study including 10 patients with idiopathic focal axillary hyperhidrosis since childhood that was unresponsive to other nonsurgical treatments, who received (BoNT-A) unilaterally and BoNT-B contralaterally. In 2011, Frasson E et al.  compared2).Each patient was injected in one axilla with 50 U of (BoNT-A) diluted with 1 mL of 0.9% sterile physiologic saline without preservatives and in the contralateral axilla with 2500 U of BoNT-B diluted with 0.5 mL of 0.9% sterile physiologic saline without preservatives. The identified hyperhidrotic area was pen-marked and subdivided into 2 × 2-cm square (4 cmThe toxin was injected in amounts of 0.025 mL (BoNT-B) and 0.050 mL (BoNT-A) intradermally. All patients received the same amount of toxins divided among the same injection points. All patients reported a reduction in axillary sweat production. Mean pretreatment sweat production rates and areas were similar bilaterally. After BoNT injections, patients responded to treatment until month six. At 1 and 2 weeks and 1, 3, and 6 months after treatment, sweat weight and area decreased significantly more in the BoNT-B side than in the BoNT-A side. Moreover, patients’ treatment satisfaction scores were significantly higher for the BoNT-B than for the BoNT-A treatment until month three, and treatment began acting earlier in the BoNT-B side compared to the A/Ona side. All patients tolerated intradermally injected BoNTs, although some experienced mild pain, especially during BoNT-B injections. No hematomas developed at the injection site, nor did any participant report systemic adverse effects. The authors provided objective evidence that BoNT-B is safe and effective for treating bilateral axillary hyperhidrosis and, when administered at the same dose ratio of 1:50 used for the motor system, it blocks sweating better than BoNT-A. ®) for palmar hyperhidrosis, through a double-blind, randomized, placebo-controlled study, including twenty patients (10 men and 10 women) diagnosed with palmar hyperhidrosis, who were injected with either Myobloc®  or a 1.0 mL vehicle  into bilateral palms (15 patients received Myobloc® and 5 received the placebo). The participants were followed until sweating returned to baseline levels. The main outcome measures were safety, efficacy versus placebo, and the duration of effect. A significant difference was found in treatment response at day 30, as determined by participant assessments, between 15 participants injected with Myobloc® and 3 participants injected with the placebo. The duration of action ranged from 2.3 to 4.9 months, with a mean duration of 3.8 months. The single most-reported adverse event was dry mouth or throat, which was reported by 18 of 20 participants. The adverse event profile also included indigestion or heartburn (60%), excessively dry hands (60%), muscle weakness (60%), and decreased grip strength (50%). The authors stated that Myobloc® proved to be effective for the treatment of palmar hyperhidrosis, and it had a rapid onset, with most participants responding within 1 week. The duration of action ranged from 2.3 to 4.9 months, with a mean of 3.8 months. The adverse event profile included dry mouth, indigestion or heartburn, excessively dry hands, muscle weakness, and decreased grip strength, as already described in the literature. Results reported in ® plus BoNT-B toxin-Neurobloc®) of palmar hyperhidrosis on patients’ quality of life. Twenty-six patients with palmar hyperhidrosis were injected with 213 ± 19 U Xeomin® and 264 ± 60 U Neurobloc® over the thenar eminences to avoid muscle weakness. At follow-up 3 weeks post-treatment, 95% of patients were satisfied, evaporation decreased >50% and the Dermatology Life Quality Index (DLQI) score significantly improved from 10.3 to 1.2. Only one patient experienced muscle weakness. In conclusion, Xeomin® in combination with Neurobloc® has an excellent effect on palmar hyperhidrosis, and Neurobloc® may be an option for use in the treatment of palmar hyperhidrosis in order to minimize muscular side effects.In 2013, Rosell K et al.  investigp < 0.001) of paper towels’ weight for the right palm at the baseline, 4, 12, and 24 weeks; and 2.8 ± 1.7, 0.5 ± 0.6, 0.8 ± 0.7, and 1.8 ± 1.25 g (p < 0.001) at same time, respectively, for the left palm. A significant reduction of mean VAS values were also detected after BoNT-B injections: 8.6 ± 1.1, 0.6 ± 0.8, 3.5 ± 2.5, and 7.1 ± 2.4 (p < 0.0001) at the baseline, 4, 12 and 24 weeks, respectively. Mild side effects consisting of local pain and hand weakness were observed in four (12.5%) subjects. The authors concluded that the use of 5000 IU BoNT-B injection in each palm was safe and significantly improved the severity of palmar hyperhidrosis.In 2014, Basciani M. et al.  reportedNo case report, case series, original prospective or retrospective studies have been reported concerning the use of BoNT-B for plantar hyperhidrosis, thus BoNT-A still represents the only serotype used for this site, with good results over the short-term, but unsatisfactory long-term results . 2/h) and gravimetry (mg/min), the time to reinjection as a measure of duration of the treatment effect and the frequency of adverse events which were captured throughout the study. BoNT-B ) was used and 5 U per injection were administered intradermally every 15 mm in a checked pattern covering the entire hyperhidrotic area, taking special care to inject the area of the forehead no closer to the eyebrows than 40 mm. Injections of hyperhidrotic areas were not limited to the face but the forehead, neck, and parts of the scalp (tonsurans) were also treated. DLQI scores were significantly improved at follow-up 2–4 weeks post-treatment and sweating was significantly reduced. Similarly, sweating before treatment monitored with transepidermal water loss and gravimetry significantly decreased after treatment. Regarding the Global Assessment of Therapy, 87% of the patients were satisfied with the treatment result. In a 2-year follow-up, 74% returned for further treatment after a median time of 5 months. Side effects were mild and the most-reported was stiffness of the forehead and the eyebrows. The authors concluded that BoNT-B seemed to be both a safe and effective treatment in craniofacial hyperhidrosis improving quality of life and reducing extreme sweating.Results reported in In 2019, Cabreus P et al.  describeCantarella G et al.  in 2010 2  injected intracutaneously in the mapped area of gustatory sweating. All patients were followed up for 12 months. One month after treatment, six of the seven patients reported that gustatory sweating and flushing had resolved, and, in the remaining patient, these symptoms had decreased. The Minor test confirmed a significant improvement. The subjective benefits remained stable for six months in four patients and for nine months in the remaining three patients; 12 months after treatment, all patients still reported some improvement. The authors concluded that BoNT-B afforded symptomatic relief in a small sample of patients with Frey’s syndrome and might be considered a potential alternative to botulinum toxin type A.Frey’s syndrome is a frequent sequela of parotidectomy, causing facial sweating and flushing because of gustatory stimuli. Although botulinum toxin type A has become a first-line therapy for Frey’s syndrome, some patients become resistant, thus the authors investigated whether another serotype, BoNT-B, might be an effective alternative. Seven patients aged 30 to 68 years, with severe Frey’s syndrome, underwent the Minor test and had 80 U of BoNT-B B per cmResults reported in ®/NeuroBloc®) at the trunk, with dosage raging from 1000 to 4000 MU. NeuroBloc®/Myobloc® 250 U/mL, was given every 15 mm with 7.5 U at each spot. The patients responded to a DLQI questionnaire before injections with BoNT-B and 3 weeks after treatment. At the follow-up visit, the participants also ranked the effect of the treatment on a five-grade scale. Three patients had residual sweating after BoNT-B treatment, and received additional anticholinergics at the follow-up visit. Those subjects eventually had a third evaluation with the DLQI. The DLQI score was, on average, 16.4 before treatment and decreased to 4.8 after BoNT-B injections. Eight out of nine patients were satisfied with the treatment. The average DLQI score decreased to 2.2 when the patients with residual sweating (n = 3) received additional anticholinergics. Adverse events from BoNT-B were mild and temporary, but dry mouth was substantial in one patient using anticholinergics. For the authors, a combination of BTX A/B and anticholinergics alleviated the hyperhidrosis with minor side-effects. We consider this treatment safe, effective, and well tolerated.A major problem in the treatment of CH is that patients typically have large areas with excessive sweating which limit the use of BoNT-A. However, BoNT-B has a greater affinity for sudomotor fibers and alleviates excessive sweating from large areas in relatively small doses ,6,20,21.Results reported in ®]) compared with a placebo in treating hyperhidrosis in the residual limbs of individuals with amputation in a randomized, double-blind, placebo-controlled pilot study. Nine male patients with eleven major amputations of the lower limbs and who complained of excessive sweating in their residual limbs were enrolled in the study between 24 September 2008 to October 28, 2011. Participants’ lower limbs were randomly assigned to receive injections of either BoNT-B (n = 7) or a placebo (n = 4). The primary efficacy variable was a minimum of 50% reduction in sweat production 4 weeks after the injection as measured via gravimetric sweat analysis after 10 min of physical exertion. Secondary analyses were performed on prosthetic function and pain. All volunteers  in the BoNT-B group achieved a minimum of 50% reduction in sweat production as compared with only 50% (two) in the placebo group. The percent reduction was significantly greater for the BoNT-B group than for the placebo group . Although both groups subjectively self-reported significant sweat reduction and improved prosthetic function (p < 0.05 for both), objective gravimetric sweat analyses significantly decreased only for the BoNT-B group . Neither group reported a change in phantom limb pain or residual limb pain (p > 0.05 for both). BoNT-B successfully reduces sweat production in individuals with residual limb hyperhidrosis, but does not affect pain. In 2006, Pasquina PF et al.  investigp = 0.027) and 3 months ; p = 0.020). Impaired quality of artificial limb use likewise improved from a baseline NRS of nine (5–9) to two (1–4) after 4 weeks and three (1–4) (p = 0.027) after 3 months, consistent with limited duration of use (p = 0.023). Steadiness of gait, quality-of-life, and work performance increased accordingly, and skin problems decreased clearly but not significantly. Unexpectedly, stump pain was also reduced ; 4 weeks: NRS, four (3–5), p = 0.109; 3 months: NRS, three (2–4), p = 0.008). For these authors, a low-dose of BoNT-B significantly reduced sweating of the residual limb, thereby improving the use of the artificial limb, steadiness of gait, and quality of life.These results are reinforced by those of Kern U et al. in 2011 . They peSince the discovery of the therapeutic potential of botulinum toxin for the treatment of strabismus in 1980, its therapeutic applications has grown dramatically over the years. Among the BoNTs family, BoNT-A represents the most well-known and well-studied, and it now provides therapy for patients in neurology, ophthalmology, pediatrics, surgery, and dermatology .® in the United States and as NeuroBloc® in Europe. A new serotype of botulinum toxin has arrived in the United States. Botulinum toxin type B (BoNT-B), known as MyoblocBotulinum Toxin Type B inhibits acetylcholine release at the neuromuscular junction via a three-stage process: (1) Heavy-chain-mediated neurospecific binding of the toxin, (2) internalization of the toxin by receptor-mediated endocytosis, and (3) ATP and pH dependent translocation of the light chain to the neuronal cytosol where it acts as a zinc-dependent endoprotease cleaving polypeptides essential for neurotransmitter release.Botulinum Toxin Type B binds to and cleaves the synaptic Vesicle Associated Membrane Protein  which is a component of the protein complex responsible for docking and fusion of the synaptic vesicle to the presynaptic membrane, a necessary step for neurotransmitter release .BoNT-A and BoNT-B have been used for a myriad of dermatologic issues in an “off-label” way . It has Axillary hyperidrosis represents the most-studied application for BoNT-B in focal hyperhidrosis, with five reported studies including 57 patients treated with BoNT-B with dosage ranging from 1000 to 5000 MU. Taken together, data on efficacy and its safety profile indicate that BoNT-B has a rapid onset of action, with most participants responding with a cessation of sweating in 5–7 days. The duration of action ranged from 2.2 to 8.1 months, with a mean of 5.0 months. The adverse event profile is generally acceptable and includes bruising and dry eye (in about 25% of subjects), dry mouth (in 35%), and flu-like symptoms (in 30%). Less studies concerning the use of BoNT-B in palmar hyperhidrosis have been reported, although the total number of patients with palmar hyperhidrosis receiving a treatment with BoNT-B was greater for palmar than axillary hyperhidrosis (78 vs. 57). The anhidrotic effect of BoNT-B largely depends on the dose used, with significant reduction of palmar sweating after injection of BTX-B at a dose of 5000 U per palm ranging from 2.3 to 4.9 months, with a mean duration of 3.8 months. The most commonly reported adverse event was dry mouth/throat, which is reported by almost 90% of the patients. The adverse effect profile also includes indigestion/heartburn (in 60%), excessively dry hands (in 60%), muscle weakness (in 60%), and decreased grip strength (in 50%). BoNT-B also had a rapid onset, with most participants responding within 1 week.More studies evaluating the use of BTX-B to manage both axillary and palmar hyperhidrosis are clearly required, mostly because there is no consensus yet on optimal dosing regiments for using BTX-B as well as conversion factors for converting BoNT-A to BoNT-B .Few data have been reported on craniofacial hyperhidrosis treatment with BoNT-B, including 53 patients (seven among them with Frey’s syndrome). The overall results indicate BoNT-B to be a safe and effective treatment in craniofacial hyperhidrosis, with no different emerging side effects, compared to axillary hyperhidrosis.With regard to compensatory hyperhidrosis, the greatest challenge consists in its treatment, as it is usually diffuse on larger areas than focal hyperhidrosis, so that BoNT-A at standardized dosage is of limited efficacy. For this reason, BoNT-B, which has a greater affinity for sudomotor fibers, could alleviate excessive sweating on large areas with relatively small doses, and preliminary data seem to confirm this specific valence of BoNT-B.Finally, preliminary data have been reported about the use of BoNT-B for treatment of residual hyperhidrosis in amputated patients. The impact of focal hyperhidrosis is tremendous in amputees, and we are in dire need for systematic, larger studies to confirm the preliminary results of the efficacy of BoNT-B. Two major concerns are related to BoNT-B use in focal hyperhidrosis: the increasing incidence in the severity of related side effects and the increased pain at injection site, compared to BoNT-A. BoNT-B has been shown to produce more systemic side effects than BoNT-A and this is indicative of a difference between BoNT-B and BoNT-A. It has been hypothesized that BoNT-B may have a higher affinity for autonomic nerve endings than BoNT-A, although the mechanism of action underlying this affinity remains only speculative. However, BoNT-B should be used with caution in patients with pre-existing autonomic dysfunction and in those with conditions for which anticholinergic therapy is contraindicated .On the other hand, the postulated preferential action of BoNT-B on the autonomic nervous system, could give BoNT-B a greater specificity of use in the management of palmar hyperhidrosis, with a hypothetical potential lower risk of generating hyposthenia, compared with BoNT-A.To verify if these assumptions are real and BoNT-B merits increasing use in palmar hyperhidrosis to guarantee a long-lasting benefit without hand muscle weakening, the increase in experience of use is mandatory. Another specific side effect that becomes more and more apparent with increasing clinical use of BoNT-B is injection site pain. The incidence of injection site pain appears to be higher for BoNT-B than for BoNT-A. The reason that BoNT-B is less comfortable than BoNT-A remains unknown, although several hypotheses have been considered; the greater volume that is injected, the concentration of active or inactive proteins, or even the lower pH of the solution which is required to stabilize the BoNT-B solution. The latter hypothesis seems to be the most accredited. Diluting the original BoNT-B preparation with saline to make 250 MU in 2.5 mL also dilutes protons below the pain threshold; it can reduce injection-related discomfort for patients. Further studies are needed to investigate both causes, as well as solutions, to this problem.A strong point in favor of the use of BoNT-B is its efficacy extended to patients resistant to BoNT-A. While the potential for secondary resistance to BoNT-B remains unclear, it may occur less than with BoNT-A as commercially available BoNT-B manufacturing procedures do not include freeze-drying and the product does not require reconstitution before use. However, switching to a BoNT-B formulation is undesirable from an immunological perspective, given that BoNT-B has a higher immunogenicity than BoNT-A ,31. Future research should increase studies comparing how BoNT-A and BoNT-B affect the autonomic and motor nervous systems and how long their action on both systems lasts.A PubMed search from 2000 to November 2022 was performed to identify any reports on the use of BoNT-B in focal hyperhidrosis. We detected these articles using the terms “botulinum toxin type B and focal hyperhidrosis”, “botulinum toxin type B and palmar hyperhidrosis”, “botulinum toxin type B and axillary hyperhidrosis”, “botulinum toxin type B and cranio-facial hyperhidrosis”, “botulinum toxin type B and plantar hyperhidrosis”, “botulinum toxin type B and compensatory hyperhidrosis”, and “botulinum toxin type B and residual hyperhidrosis”. Only studies in English were reviewed. All studies that met the criteria were included and summarized in this review. The PRISMA study flowchart is shown in


Highly functioning adults with a sedentary lifestyle benefit physically and mentally from forest therapy and mountain hiking.Women predominantly benefited highly from mountain hiking regarding hemopoietic system and aerobic capacity. Both genders profited mentally from contact with nature.What are the main findings?Forest therapy and mountain hiking could be safe and health-promoting interventions for high-functioning adults with a sedentary lifestyle and could be applied in primary prevention as well as in secondary prevention.What is the implication of the main finding?Background: Lifelong physical activity is related to longer health span, which is reflected at an individual level, and is of substantial socioeconomic relevance. Sedentary lifestyles, on the other hand, pose an increasingly major public health problem. In addition, the COVID-19 pandemic had a negative impact on activity levels and well-being. Previous research indicates that contact with nature might improve exercise levels as well as well-being. Methods: This randomized, controlled clinical trial (ANKER-study) investigated the effects of two types of nature-based therapies (forest therapy and mountain hiking) in couples  with a sedentary or inactive lifestyle on health-related quality of life, relationship quality and other psychological and physiological parameters. Results: The results of this study displayed that healthy and highly functioning women and men with sedentary lifestyles mentally benefit from contact with nature . The gender-specific effect on women is most visible in the physiological outcomes  of mountain hiking. Men and women showed small improvements in blood pressure as a result of the interventions. Conclusions: The ANKER-study provides a method for valid comparison of forest therapy interventions for the first time. Regarding the COVID-19 pandemic, the nature-based intervention presented could offer a multimodal contribution to maintaining a more active lifestyle, further contact with nature that affects peoples physical as well as mental health, and an improvement in social interaction. There is a considerable body of evidence on sustaining health benefits of being active and a scientific consensus that physical activity—in adequate dosage—should be used in the promotion, maintenance and recovering of physical as well as mental health ,2,3. LifIn order to promote the enjoyment of physical activity and, thus, encourage adherence and support a healthy lifespan for all individuals, it is important to consider different lifestyles . ParticuClinically relevant health benefits for a wide range of conditions could be achieved at levels of physical activity clearly below current international recommendations  . However.Therefore, effective lifestyle interventions should take an integrated approach to health and promote the importance of healthy lifestyles such as reducing sedentary time and increasing physical activity, additionally promoting healthy eating as well as managing stress . FurtherFurther, evidence shows that exposure to immunoregulatory microorganisms (“Old Friends”) is essential for humans to decrease immune (re)activity and inflammation ,18. FollAdditionally, predominantly in high-income countries, a significant decline in physical activity has been observed . Globally, more than a quarter of the population has insufficient levels of physical activity. Furthermore, beyond reduced contact with nature, urbanization thus leads to lifestyle changes as work and leisure activities shift indoors, encouraging inactive or sedentary behavior . These cA possible approach to reconnect with nature and “Old Friends” as well as to increase physical activity, could be Green Exercise. This means exercise in nature, as it is the case in particular with mountain hiking or, in a less intensive form, with forest therapy. Mountain hiking can be described as a long-lasting activity of moderate intensity at different altitudes . This leLikewise, the positive effect of nature and its specific elements on human health is the essential component of forest therapy: This approach of nature connection therapy, also known as “forest bathing”/”Shinrin-Yoku”, is a collective term for activities designed to improve human health or well-being in a forest environment. The crucial element of forest therapy is recognition for the forest environment using the five human senses, which can be combined with meditation, breathing exercises, forest walks, various recreational activities, and cognitive behavioral therapy . AdditioForest therapy has been shown to be effective in improving immune function; a hypothesis for this is that the forest biodiversity and microbiome has a significant impact; more precisely, phytoncides from trees can reverse stress-induced immunosuppression and thus, normalize immune function and neuroendocrine hormone levels , which mAlthough, there are further studies on positive health effects of forests ,37,38, mThe purpose of the ANKER-study (“Algund Nature and Climate Therapy: Green Exercise vs. Nature Connection”) is to investigate long-term effects of two types of nature-based therapies—mountain hiking vs. forest therapy—in couples with a sedentary lifestyle on health-related quality of life (HRQOL), relationship quality and other psychological and physiological parameters.https://doi.org/10.3390/ijerph19073888, accessed on 1 October 2022 [The present data were collected as part of the ANKER-Study (ISRCTN43292449); this randomized, controlled clinical trial investigated the effects of two types of nature-based therapies in couples with a sedentary lifestyle on health-related quality of life, quality of relationship, psychological and physiological parameters. Therefore, two intervention groups—a hiking group (HG) and a forest therapy group (FTG)—were included. The allocation ratio was set at equal sample size. The study protocol was approved by the Ethics Committee of Bolzano, Italy (reference number 18-2019) and can be accessed with further details on materials and methods at ber 2022 . https://www.klimatherapie.eu/, accessed on 1 October 2022) and advertisement on social media channels between April 2019 and May 2021 allover Austria and Germany. Due to limited human resources, the recruitment and, also, the intervention periods had to be extended as described, as scientific staff performed the extensive measurements on site. The seven-day vacation with mountain hiking and forest therapy in South Tyrol/Italy itself was the incentive to participate.Couples with a sedentary lifestyle were the defined group of interest in the ANKER-Study, therefore, participants had to meet the following inclusion criteria: age 50–60 years, relationship-duration > 1 year, BMI ≥ 25 and ≤30, sedentary lifestyle  < 3.00 METmin/week) and the ability to participate in moderate hiking tours . The following exclusion criteria were applied: active lifestyle, immunologically mediated chronic conditions or immunodeficiency, severe respiratory diseases, acute or untreated psychiatric disorders, uncontrolled hypertension, uncontrolled metabolic disease, acute infection or fever, diagnosis of or treatment for malignant neoplastic disorders within the last 5 years, arteriosclerotic event < 6 months before enrollment, cardiac insufficiency, renal insufficiency, diagnosis or history of alcoholism, current recreational drug use, currently smoking > 10 cigarettes/day, orthopedic contraindications for hiking, medication intake > 5mg/d prednisone, colchicine, Imuran, methotrexate, azathioprine, cyclophosphamide or cyclosporine, intake of weight-loss drugs or preparations as well as pregnancy. By selecting people between 50 and 60 years of age, the interventions studied could contribute to maintaining the ability to work in this age group, as this would be a primary preventive measure to maintain health. The participating couples were recruited online via webpage , a region that is characterized by its mild, nearly Mediterranean climate. All participants were housed in local hotels, got the same meals, and did not receive lifestyle recommendations during the non-intervention phase.The HG took part in guided, moderate hiking tours every day (3–4 h), except for one rest day in the middle of the intervention week. The nature group received daily 3–4 h of standardized nature connection therapy sessions with a psychologist trained for this purpose. Forest therapy was characterized by low physical activity and oriented on a holistic framework that promoted meaningful connections at the three levels of “connection with nature”, “connection with others”, and “connection with oneself”. In forest therapy, each day had a specific theme that was worked on . After a practice phase, the participants finished the intervention each day with a written self-reflection. These interventions were conducted in two separate, but identical sequences: The first part of the study population completed the ANKER-Study in October 2019, the second part in June 2021.Medical examinations at T1 , and T2  were performed at the Department of Sports Medicine, Tappeiner Hospital, Merano . Follow-up examinations at T3 (day 60) took place at the Paracelsus Medical University, Salzburg (Austria); the follow-up examination at T4 (day 180) was carried out as an online survey. More details are shown in The ANKER study investigated the following long-term effects on T1, T2, T3 and T4: Health-related quality of life , EuroQol (EQ-5D-5L), Quality of relationship (Partnership Questionnaire (PFB) and Problem List (PL)) as primary outcomes. A further primary outcome—the Intercultural Quality of Life Comic will become part of a separate paper due to its extensive results and an additional validation (throughout the following questionnaires: Subjective impairments of the participants (Complaints list (B-L’), BFI-10 questionnaire (10 Item Big Five Inventory) and Health-related control beliefs (HLOC) in this context. ®), aerobic fitness (Chester step test) and transepithelial water loss . Additionally, anthropometric measures  were acquired. The motion profile of the heart rate monitors (watches on the wrists of the subjects) was obtained over the time of the intervention. Secondary parameters were nature connectedness , Nature Relatedness Scale (NRS) and Inclusion to Nature Scale (INS)), sociopsychological well-being in the sense of flourishing of personality , life satisfaction , health-related control beliefs (FEGK), subjective impairments of the participants Mindfulness . Measurements of the International Physical Activity Questionnaire Short Form (IPAQ-SF) were collected at T1, T3 and T4 to assess a possible long-term impact of the intervention on the subjects’ physical activity behavior. Additionally, the following physiological parameters were collected on T1, T2 and T3: static balance (MFT-S3 Check), body composition via Bio Impedance Analysis , differential blood count (Forearm venous blood), fractional exhaled nitric oxide . Statistical significance was set at the level of α < 0.05. Baseline data were analyzed using unpaired Students-Fully nonparametric variance type tests (ANOVA type statistics) from the nparLD  package  were usePairwise randomization of the couples to groups was completed according to the following stratification factors: Age, general health (PHQ-9), closeness to nature (NRS-6), BMI, activity level (IPAQ-SF) and relationship duration . Due to Two hundred and fifty-seven individuals were screened for eligibility to participate in this study, 165 were excluded: 49 individuals did not meet the inclusion criteria, 116 individuals declined to participate due to personal reasons or the COVID-19 pandemic. In October 2019, 24 people were enrolled in the HG and 28 in the FTG; in June 2021, 20 people were enrolled in the HG and 18 in the FTG. This time gap occurred due to limited scientific staff resources. The four groups were pooled into one HG (n = 42) and one FTG (n = 46) for examination. Two participants from the HG did not receive allocated intervention  and two further participants were excluded from the analysis because they did not meet inclusion criteria anymore (relationship status). Consequently, 42 participants of the HG and 46 participants of the forest therapy group  were incp = 0.06, W = 742). An extended overview of all other baseline characteristics is summarized in Except for relationship duration, baseline characteristics showed no relevant differences in inclusion criteria between the study groups . In the The hiking intervention as well as the forest therapy were well tolerated by all participants. No adverse effects or harms were observed in any group. On average, the mountain hikes lasted 03:29 h, the participants covered a distance of 7.36 km and 521 m of altitude. The FTG participated each day in standardized forest therapy sessions for 3–4 h, assisted by a psychologist. The sessions were characterized by low physical activity . Both inResults from the F1-LD-F1 analysis of primary outcomes are accessible in p < 0.01). Post hoc tests did not show any interaction effects at the single time points, indicating a parallel improvement in both groups. The SF-12 Physical Component was rated significantly higher by the HG at baseline than by the FTG . Significant main effects for group (p < 0.01) and time (p < 0.01) were observed. Post hoc test did not show any interaction effects at the single time points. The parallel RTE-profile in combination with the significant baseline difference indicates again a parallel improvement in both groups. For the SF-12 Mental Component, a significant main effect for time was found (p = 0.02), indicating a parallel development in both groups. The F1-LD-F1 model for the SF-12 total score revealed a significant main effect for time (p < 0.01). Post hoc test showed no interaction effects at the single time points, thus indicating a parallel improvement in both groups. The EQ5D-5L Index presented with a significant main effect for group (p = 0.02) and time (p < 0.01). A post hoc test failed to reveal any interaction effects at the single time points, thus indicating together with the RTE profile a parallel improvement in both groups until day 60. The improvement continues in the HG, whereas the EQ-5D-5L index decreases in the FTG towards day 180.The F1-LD-F1 model for the EQ5D-5L visual analogue scale revealed a significant main effect for time (p < 0.01). Post hoc test revealed no interaction effects at the single time points. The relatively flat RTE profiles indicate, that the quality of relationship did not change in any group throughout the study. The F1-LD-F1 model for the subscale happiness revealed a significant main effect for time (p < 0.01) and a trend for group*time (p = 0.07). Post hoc test failed to identify any interaction effects. Again, the relatively flat RTE-profile indicates, that the happiness subscale did not change throughout the study. A significant main effect for time (p = 0.02) and for group*time (p = 0.03) was identified for the problem list (PL). No interaction effects at the single time points were found. However, the participants in both groups rated at all four time points only 1–2 relationship areas as ≥ 2.Regarding the quality of relationship , both groups start with high baseline levels . Scores ≥ 54 are considered as a mostly happy relationship. The F1-LD-F1 model for the PFB Total Score revealed a significant main effect for time (p < 0.01) and hematocrit (p < 0.01). p < 0.01) and group*time (p < 0.01) in the female subgroup. Post hoc test indicates a significant decrease in erythrocytes in the female HG towards day 7 (p = 0.04) with a return to baseline at day 60 (p = 0.08). Within the male subgroup, only a significant main effect for time (p < 0.01) was found. Post hoc test did not show any interaction effects. However, the RTE-profile also indicates a decrease in erythrocytes in the male HG.The F1-LD-F1 model for erythrocytes identified a significant main effect for time (p < 0.01) and group*time (p = 0.03) was identified for hematocrit. The male subgroup presents with a significant main effect for group (p = 0.02) and time (p < 0.01). Post hoc test and RTE profile indicate a decrease in hematocrit for both sexes in the HG towards day 7 with a return to baseline at day 60.Within the female subgroup, a significant main effect for time (p < 0.01) indicated an increase towards day 7 and a return to baseline at day 60 in both groups. However, the immature reticulocyte fraction (IRF) presented with a significant main effect for time (p < 0.01) and group*time (p = 0.05). Post hoc test and RTE profile indicated an increase in the HG towards day 7 (p = 0.07) and a return to baseline at day 60.In contrast, the F1-LD-F1 model for reticulocytes showed an opposite dynamic. A main effect for time (p < 0.01), indicating anti-inflammatory effects of both interventions. Post hoc test did not show any interaction effects at the single time points. Statistical analysis of the white blood cell count revealed a significant main effect for time (p = 0.04) and group*time (p = 0.03). Post hoc test found a significant interaction effect at day 7 (p < 0.01) indicating an improvement in the HG. Within the male subgroup, a significant main effect for time (p = 0.04) was found. Post hoc test failed to show any interaction effects at the single time points. However, the RTE profile indicates an improvement in aerobic capacity in the male FTG. a graphical representation, see F1-LD-F1 model for aerobic capacity in the female subgroup revealed a significant main effect for time (p < 0.01) indicates an improvement in both groups. However, the improvement is in the HG (T2-T1: 1.16%) and FTG (T2-T1: 2.63%) rather small. In contrast, the sensorimotor index did not show any significant changes. Participants in both groups already reach around 77% in the S3-Check Stability index at the beginning of the study. A significant main effect for time (p < 0.01) indicates a decrease in systolic blood pressure in both groups . Diastolic blood pressure presented also with a significant main effect for time (p = 0.02), indicating an improvement . F1-LD-F1 models for pulse and SpO2 revealed significant main effects for time (p < 0.01), but post hoc tests failed to show interaction effects at the single time points. No significant group-, time- or interaction effects for skin parameters  were observed. In F1-LD-F1 models for blood pressure found similar effects for systolic and diastolic blood pressure. A significant main effect for time (p < 0.01), reactance (p < 0.01), phase angle (p < 0.01), weight (p < 0.01), fat mass index (p < 0.01), fat free mass index (p < 0.01), skeletal muscle mass index (p < 0.01), body cell mass index (p < 0.01) and hydration (p < 0.01).Results from the F1-LD-F1 analysis of Body Parameters are presented in p < 0.01) and group*time (p = 0.05). Post hoc test did not show any interaction effects. Although a significant time effect was found, the changes in BMI are rather small and lack any clinical relevance. Additionally, body weight presents in both sexes with a significant main effect for time  but lacking any clinical relevance. F1-LD-F1 model for body mass index revealed a significant main effect for time (p < 0.01) and group*time (p = 0.01) in the female subgroup. The male subgroup presented only with a main effect for time (p < 0.01). F1-LD-F1 models for resistance found for both subgroups a significant main effect for time . No significant effects were found for phase angle in any group. F1-LD-F1 model for reactance revealed a significant main effect for time (p < 0.01) and group*time (p = 0.03), indicating an increase in fat free mass in the HG. Furthermore, the Skeletal Muscles Index revealed also a significant main effect for time (p < 0.01) and group*time (p = 0.01) in the female subgroup, indicating an increase in skeletal muscle mass in the HG. The F1-LD-F1 model for hydration presented with a significant main effect for time (p = 0.02), indicating an increase in hydration in the HG. No effects were found for fat mass index.Within the female subgroup, the Fat Free Mass Index presented with a significant main effect for time (p < 0.01) and Skeletal Muscles Index (p < 0.01). No effects were found for hydration and fat mass index. Within the male subgroup, significant main effects for time were found for Fat Free Mass Index (p = 0.01) and flourishing scale . For all these questionnaires, it should be noted that the participants already had high scores initially and consequently, the changes in the scores were minor.F2-LD-F1 models revealed gender-specific effects for the connectedness to nature scale , indicating a parallel improvement in both groups. Internal health-related control beliefs  revealed a significant main effect for group (p = 0.01) and time (p = 0.02). Post hoc test did not find any interaction effects at the single time points. External health-related control beliefs  presented with a main effect for time (p = 0.01). Within the female subgroup, the flourishing scale revealed a significant main effect for time (p = 0.01) indicating together with the RTE-profile a personal flourishing in the HG. No significant effects were found for the male subgroup. F1-LD-F1 models for the IPAQ-SF revealed significant main effects for time, indicating a parallel development in both groups. The physical activity increased in both groups slightly throughout the study. For further details, all questionnaire data can be found in Nature connectedness increased in both intervention groups as significant main effects for time were found for the CNS, INS and NRS-6. F1-LD-F1 model for MAAS Total revealed only a significant main effect for time , which already approached the recommended range immediately after the one-week intervention , so a clinically relevant change can be assumed. Gatterer et al. stated that in elderly persons with hypertension, moderate intensity hiking only at weekends may reduce systolic blood pressure; otherwise, it did not improve cardiovascular risk factors in healthy older people. They did not find gender-specific differences . SimilarForest therapy in the ANKER-study was professionally guided by a specialized psychologist. Findings exhibited that guided forest therapy programs promote positive emotional changes and social bonding through interaction with others . Which, Moreover, there are also differences in the gender-specific influence of COVID-19: Amplified risk for developing mental health problems for women during the pandemic is experienced with pregnancy or intimate partner violence . In contDuring the COVID-19 pandemic, concerned women, older persons, college-educated respondents, and white-collar workers noted the greatest use of the forest as a recreation site. Nonetheless, women and people with elementary education expressed the most fears related to visiting the forest. In contrast, men and people who lived in rural areas and small towns, as well as respondents who performed activities related to the forest, were most involved in exploring and working in the forest and . Other fhttps://doi.org/10.3390/ijerph19073888 access on 1 October 2022). In this context, it would be of interest to know what influence the forest characteristics themselves have on the results, here further research and comparisons with other forests are needed. The ANKER-study was conducted in a small forest area, with mixed, medium old and rather high stock of trees in a quiet environment [To enhance the methodological quality and evidence, while decreasing the risk of bias, as required in reviews on the topic of forest therapy ,38,64, tironment .Nature relatedness in the ANKER-Study was assessed via three questionnaires . The results indicated that connectedness to nature is a stable personality characteristic, as the scores changed only slightly, even though significance occurred over time, i.e., low clinical significance can be expected. Furthermore, it could be assumed that there was a selection bias in the present study population, because especially persons with a high affinity to nature would volunteer to participate in such a study design, which is also reflected in the high, stable values of these three questionnaires. As the bootstrap sample size simulation based on the results of the ANKER study shows, a sample size of n ≥ 80 persons is recommended in order to similarly match the psychological and physiological parameters of nature-based interventions. Although, in this study only about half of the recommended subjects participated, significant results could be obtained already, therefore higher case numbers are likely to generate even clearer and clinically more relevant results. Additionally, further research specific to women with regard to mountain hiking and also forest therapy should be conducted to further elucidate the effects of these interventions on female gender and to ascertain their backgrounds, e.g., fears related to visiting the forest . FurtherBuckley and Brough  pointed The ANKER-study, with its forest classification survey, provides for the first time a method for a more valid comparison of forest therapy interventions. The long-term results of this study showed that healthy, highly functioning women and men with sedentary lifestyles benefit mentally from contact with nature. The gender-specific effect of mountain hiking on women is most visible in physiological parameters. Further research should investigate forest therapy as a health-promoting intervention for individuals with immobility or internal diseases. Advanced gender-specific research, especially with regard to these interventions on women, is also recommended. Regarding the COVID-19 pandemic, the nature-based intervention presented could offer a multimodal contribution to maintaining or creating a more active lifestyle, further contact with nature that affects people’s physical as well as mental health, and an improvement in social interaction. Overall, focus should be on high quality research to make these types of nature-based therapies more applicable and prescribable for a working-adult population.